Comparison of in vivo bioactivity and compressive strength of a novel superporous hydroxyapatite with beta-tricalcium phosphates.

INTRODUCTION: Superporous hydroxyapatite (HAp-S) is a novel bone substitute that contains three-dimensionally interconnected macropores with micropores, which stimulate bone ingrowth into the material. METHOD: We investigated the in vivo behaviour of HAp-S by comparing its bioactivity and biomechanical properties with beta-tricalcium phosphates (ß-TCP). HAp-S or ß-TCP was implanted in the lateral femoral condyle of rabbits. In vivo bioactivity of each material, including bone ingrowth and material resorption, was quantitatively evaluated by micro-CT and the ultimate compressive strength of the bone-material composite was also measured. Micro-CT showed that bone ingrowth in the HAp-S group significantly increased over time, while no significant increase was observed after 8 weeks in the ß-TCP group. RESULTS: Although both materials showed gradual material resorption, ß-TCP resorption was significantly greater than HAp-S. The ultimate compressive strength in the HAp-S group significantly increased over time up to six times its original value, while there was no significant increase in the ß-TCP group. These results show that HAp-S resorption is concurrent with bone ingrowth, resulting in increasing compressive strength over 12 weeks. On the other hand, ß-TCP resorption is fast but unaccompanied by bone ingrowth; consequently, it remains relatively fragile at least in the early period after implantation. Although these highly porous materials themselves are structurally and mechanically similar, there are significant differences in in vivo behaviour depending on the material composition. CONCLUSION: These findings should be kept in mind when choosing the highly porous ceramics.

Long-term antibacterial activity of vancomycin from calcium phosphate cement in vivo.

BACKGROUND: Periprosthetic joint infection is a major complication of total joint arthroplasty, with treatment requiring a two-stage exchange procedure and 6 weeks of systemic antibiotics. However, depending on the infection site, intravenous delivery of antibiotics like vancomycin (VCM) can have poor tissue transferability, thus reducing their therapeutic effect. OBJECTIVE: This study demonstrates the 24-week in vivo release profile and antibacterial activity of VCM from calcium phosphate cement impregnated with VCM (CPC/VCM) and compares them with those from polymethylmethacrylate impregnated with VCM (PMMA/VCM). METHODS: Rats were implanted with the test specimens between the fascia and quadriceps. After implantation for 24 weeks, the test specimens were removed and residual VCM was extracted to calculate the concentration of VCM released into rat tissues. We also examined the antibacterial activity of releasable VCM from the removed test specimens by placing them directly onto the surface of agar. RESULTS: CPC/VCM released greater concentrations of VCM for a longer period of time within the 24 weeks than PMMA/VCM. Moreover, CPC/VCM released 1.4 to 26.1-fold more VCM than PMMA/VCM. Using Staphylococcus aureus, antibacterial activity was logarithmically correlated with VCM concentration across the entire concentration range tested (12.5-800 µg/mL). While the area within which inhibition was observed-the inhibition zone-for both CPC/VCM and PMMA/VCM formed and gradually shrank with time after implantation, that for CPC/VCM was significantly larger than that for PMMA/VCM in each week after implantation. CONCLUSION: CPC/VCM releases greater amounts of VCM with antibacterial activity for longer periods of time than PMMA/VCM, suggesting that CPC is effective for facilitating the release of antibiotics for local action in patients with established postoperative infection.

Development of superporous hydroxyapatites and their examination with a culture of primary rat osteoblasts.

When the usage of hydroxyapatite (HAp) was first approved at clinics by the Kouseishou (Japanese FDA) as a bone substitute (APACERAM), the upper limit of pore content was set at 60%. Cells play an important role in bone repair, especially in regeneration therapy, but on using these HAps, the cells cannot penetrate deeply into them because their inside pores rarely connect. To promote cell penetration into the inside of the HAps, we have developed superporous HAps (HAp-Ss). First, phosphoric acid was added to a calcium hydroxide solution, and the mixture was dried by the spray-dry method to produce fine primary particles. Then, two kinds of surfactants were used to form a large amount of pores. These two HAp-Ss have 85% porosity and interconnected pores in the inside. They were tested with a culture of primary rat osteoblasts, which showed good penetration therein. The penetrated osteoblasts maintained high alkaline phosphatase activity during the culture period. This indicates that the developed HAp-Ss are very good bone substitutes and also useful scaffolds in bone regeneration therapy.

Development of calcium phosphate cement using chitosan and citric acid for bone substitute materials.

We developed a calcium phosphate cement that could be molded into any desired shape due to its chewing-gum-like consistency after mixing. The powder component of the cement consists of alpha-tricalcium phosphate and tetracalcium phosphate, which were made by decomposition of hydroxyapatite ceramic blocks. The liquid component consists of citric acid, chitosan and glucose solution. In this study, we used 20% citric acid (group 20) and 45% citric acid (group 45). The mechanical properties and biocompatibility of this new cement were investigated. The setting times of cements were 5.5 min, in group 20 and 6.4 min, in group 45. When incubated in physiological saline, the cements were transformed to hydroxyapatite at 3, and 6 weeks, the compressive strengths were 15.6 and 20.7 MPa, in group 45 and group 20, respectively. The inflammatory response around the cement implanted on the bone and in the subcutaneous tissue in rats was more prominent in group 45 than in group 20 at 1 week after surgery. After 4 weeks, the inflammation disappeared and the cement had bound to bone in both groups. These results indicate that this new calcium phosphate cement is a suitable bone substitute material and that the concentration of citric acid in the liquid component affects its mechanical properties and biocompatibility.

Combination of porous hydroxyapatite and cationic liposomes as a vector for BMP-2 gene therapy.

The clinical significance of hydroxyapatite (HAP) as a bone substitute has become apparent in recent years and bone morphogenetic protein (BMP) a substance which induces bone has attracted much attention. In this study, a 1.2 cm diameter bone defects created on rabbit cranium were treated with the BMP-2 gene (cDNA plasmid) introduced with porous HAP after completion of hemostasis and the resultant bone formation was analyzed histopathologically. The amounts of bone formation was compared BMP-2 cDNA plasmids were not combined with cationic liposomes as a vector. Four groups of rabbits were compared. In the HAP group the cranial bone defect was treated with HAP containing 40 microg of liposomes and a dummy gene (PU). The BMP gene HAP group was treated with HAP soaked in liposomes and 10 microg of the BMP-2 gene. In addition, a group was treated with the gene without implanting HAP. Bone formation on the cranial defects was evaluated 3, 6 and 9 weeks after the operation, by X-ray and histopathological examinations. Three weeks after the operation there was vigorous bone formation in the cranial defect in the group which received the BMP-2 gene without HAP, and complete ossification was observed at 9 weeks. In the group which received HAP containing the BMP-2 gene, although new bone formation was evident surrounding the scaffold 3 weeks post-operation, the induced bone tissue did not fill all the pores of the scaffold even at 9 weeks post-operation. These results confirm the clinical usefulness of gene therapy for bone formation, using the BMP-2 gene combined with cationic liposomes as a vector. It is possible that the effects of administering the BMP-2 gene will be improved by specializing the microstructure of scaffold for gene therapy.

Tissue response to a newly developed calcium phosphate cement containing succinic acid and carboxymethyl-chitin.

We developed a new calcium phosphate cement containing succinic acid and carboxymethyl-chitin in the liquid component. In this study, the biocompatibility and osteoconductivity of this new cement were investigated. After mixing, cement in putty form was implanted immediately between the periosteum and parietal bone and in the subcutaneous tissues of rats. In control cement, distilled water was used instead of the liquid component. In addition to histological evaluations, analyses with X-ray diffraction and Fourier transform infrared were performed for the subcutaneously implanted cements. Histological examination showed slight inflammation around the new cement on the bone and in the subcutaneous tissue at 1 week after surgery. At 2 weeks, the cement was partially bound to the parietal bone. The extent of the surface of the new cement directly in contact with the bone increased with time, and most of the undersurface of the new cement bound to the host parietal bone by 8 weeks. Analysis by X-ray diffraction showed that the new cement in the subcutaneous tissue was transformed into hydroxyapatite by 8 weeks. These results indicate that this new calcium phosphate cement is useful as a bone substitute material.